Toward gene therapy for growth hormone deficiency via salivary gland expression of growth hormone

Abstract

Objectives Salivary glands are useful targets for gene therapeutics. After gene transfer into salivary glands, regulated secretory pathway proteins, such as human growth hormone, are secreted into saliva, whereas constitutive secretory pathway proteins, such as erythropoietin, are secreted into the bloodstream. Secretion of human growth hormone into the saliva is not therapeutically useful. In this study, we attempted to re-direct the secretion of transgenic human growth hormone from the saliva to the serum by site-directed mutagenesis. Materials and Methods We tested human growth hormone mutants first in vitro with AtT20 cells, a model endocrine cell line that exhibits polarized secretion of regulated secretory pathway proteins. Selected mutants were further studied in vivo using adenoviral-mediated gene transfer to rat submandibular glands. Results We identified two mutants with differences in secretion behaviour compared to wild type human growth hormone. One mutant, ΔN1-6, was detected in the serum of transduced rats demonstrating that expression of this mutant in the salivary gland resulted in its secretion through the constitutive secretory pathway. Conclusion This study demonstrates that mutagenesis of therapeutic proteins normally destined for the regulated secretory pathway may result in their secretion via the constitutive secretory pathway into the circulation for potential therapeutic benefit.

Potentialities of Gene Therapy in Pediatric Endocrinology

“…Besides skeletal muscle cells, other genetically modified cell types have been used as GH delivery devices in animal models, such as fibroblasts [51, 52], bone marrow stromal cells [53], and salivary glands [54]. Recently, Higuti et al [55] investigated a gene therapy approach based on the injection, in young and old lit/scid dwarf mice, of a plasmid in tibialis cranialis muscle encoding the hGH under the control of ubiquitin C promoter, followed by electrotransfer.…”

Secretions of Human Salivary Gland

“…Gene therapy has been developed to deliver growth hormone in deficiency states by salivary gland expression of growth hormone [16].…”

Advances in salivary gland gene therapy – oral and systemic implications

“…There are only a few reports of small incremental steps being published since 2013, e.g., [64–67]. More importantly, there are no applications of which we are aware that are close to being considered for clinical testing.…”

“…Thus, it seemed that the C-terminal POMC-like motif represented a probable sorting signal for hGH entry or retention in the forming secretory granules of the RSP. Over an approximately 10-year period, we focused on mutating the comparable motif in hGH with minimal success, with success being defined as significantly increasing the proportion of synthesized transgenic hGH secreted into the bloodstream [67, 75]. Thus, despite multiple studies from several laboratories showing that potentially therapeutic transgenic proteins can be secreted from salivary glands in an endocrine manner, it is currently impossible to predict how such proteins will be sorted following gene transfer to human salivary glands.…”